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Research Detail

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M. M. Khatun
National Institute of Biotechnology, Ganakbari, Savar, Dhaka-1349, Bangladesh

T. Tanny
National Institute of Biotechnology, Ganakbari, Savar, Dhaka-1349, Bangladesh

Abdur. M. Razzak
Pohang University of Science and Technology, Pohang 790–784, South Korea

Firoz Alam M
Department of Biotechnology and Genetic Engineering, Islamic University, Kushtia, Bangladesh

Ekhlas Uddin M
Department of Biotechnology and Genetic Engineering, Islamic University, Kushtia, Bangladesh

Ruhul Amin
Department of Biotechnology and Genetic Engineering, Islamic University, Kushtia, Bangladesh

S. Yesmin
National Institute of Biotechnology, Ganakbari, Savar, Dhaka-1349, Bangladesh

Ginger is one of the most important spices crop in all over the world. But, inadequate multiplication rate and disease transmittance are hampers ginger cultivation to meet the demand of high quality planting material for commercial cultivation. Therefore, an efficient in vitro regeneration protocol considers a best alternative to overcome this problem. During micropropagation surface sterilization is the most important step in preparation of explants. The effect of four different sterilizing agents: Mercuric (II) chloride (HgCl2), Sodium hypochlorite (NaOCl), Hydrogen peroxide (H2O2) and Bavistin, were evaluated for sterilization of rhizome buds of Zingiber officinale Rosc. by varying their concentration and time of exposure. The percentage of contamination, tissue damage and survival of cultures were observed. The result showed that among all sterilization treatments 0.1% Mercuric (II) chloride (HgCl2) was the most effective treatment. Highest rate of contamination free culture 86.66±0% was achieved with 0.1% (HgCl2) for 15 minutes. Whereas, sterilization with Hydrogen peroxide, Sodium hypochlorite, Bavistin were not satisfactory.

  Ginger, Contamination, Surface sterilization, Rhizome buds
  Plant Biotechnology Division of the National Institute of Biotechnology, Dhaka, Bangladesh
  
  
  Variety and Species
  Ginger

To standardize the sterilization method for explants of Zingiber officinale Rosc for micropropagation, using different types of sterilizing agents by varying their concentration and duration of exposure.

Explants and nutrient medium: The experiment was conducted at the Plant Biotechnology Division of the National Institute of Biotechnology, Dhaka, Bangladesh, with the objective to evaluate the effect of different sterilants on explants of Zingiber officinale Rosc. (Ginger) in conditions of in vitro culture. BARI ada-1 (ginger) was used in this experiment. Healthy rhizomes were kept in sand for sprouting. The stored sprouted rhizomes were used to get explants. After sterilization by using different sterilants in an autoclaved beaker or conical flask, with different concentrations and duration, rhizome buds about 1 to 2 cm long were placed in a coffee jar, test tube, or conical flask on MS medium (Duchefa, The Netherlands; Murashige and Skoog, 1962) containing 3% sucrose (Merck, Germany), solidified with 0.8% agar (BDH Chemicals Ltd., England). The pH (Jenway 3520 pH Meter, Bibby Scientific Ltd., UK) of the medium was adjusted to 5.8 before autoclaving (ALP Co. Ltd., CL-40M, Japan) at 121°C and 100 kPa for 20 min and gelling with agar.

Explant sterilization: The sprouted rhizome buds were collected in beaker or conical flask and kept under running tap water prior to sterilization in the laminar airflow cabinet. For the experiment four different kinds of sterilizing agent’s viz., Mercuric (II) chloride (HgCl2), Sodium hypochlorite (NaOCl), Hydrogen peroxide (H2O2) and Bavistin, were tested for explant sterilization by vary ing their concentration and time of exposure.

Inoculation: MS basal medium supplemented with cytokinin (0.3mg/l BAP) and auxin (0.5mg/l NAA) were used for inoculation and subsequent micropropagation of Zingiber officinale Rosc. Medium was checked for the contamination before inoculation. Sterilized explants were trimmed suitably to remove sterilizing agent affected parts/brown parts. Explants were then inoculated on the appropriate medium and labeled properly. Observation were recorded regularly till to 30 days for the contamination, tissue damage and survival cultures.

Incubation: The culture were placed in culture growth room .The growth room for maintenance of in vitro cultures had 25±2°C temperature and 60 to 70% relative humidity , with a photoperiod of 16h day light and 8h dark. Illumination was provided with incandescent lamps (50 W, Philips Agro-Lite).

Statistical Analysis: Statistical analysis was done to find out the effect of different sterilizing agents its concentration and time of exposure on the asepsis of th e said plant species. For each experiment, 12 rhizome buds and conducted at least twice. The mean values and standard deviations of contamination, tissue damage and survival cultures were calculated using computer soft ware (Microsoft Office Excel Worksheet).

  International Journal of Applied Biology and Pharmaceutical Technology [IJABPT], Volume-7(1):131-137, Jan-Mar-2016, ISSN: 0976-4550.
  http://www.ijabpt.com/pdf/32012-M.%20M.%20Khatun.pdf
Funding Source:
1.   Budget:  
  

In conclusion, among all the treatment used in this experiment, 0.1% Mercuric (II) chloride for 15 minutes found better for controlling contamination and tissue damage of Zingiber officinale Rosc. It can be describe that requirements for sterilization are different and depend on the tissue type, age and the nature of the explant used for micropropagation. However, to achieve better understanding of the actual mechanism, further research is needed to exploit the development of quick regeneration and transformation protocol for ginger.

  Journal
  


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